Integrated transcriptomic and co-expression network analysis identifies TaBI-1 as a key contributor to heat tolerance in wheat

Abstract Key message This study integrated RNA-seq and WGCNA to identify TaBI-1 as a key thermotolerance gene in wheat, with preliminary functional validation through yeast assays and nano-mediated transient expression. Heat stress severely limits global wheat production. To identify key thermotolerance genes, RNA-seq was performed on wheat flag leaves and grains at the grain-filling stage under 37 ℃ treatment with five time points (0, 0.5h, 1 h, 2 h and 4 h). Differential expression analysis revealed 16,228 and 5,480 differentially expressed genes (DEGs) in flag leaves and grains, respectively. Weighted gene co-expression network analysis (WGCNA) identified tissue-specific modules correlated with early heat stress (0.5h and 1 h), yielding 922 and 454 core genes, including the previously reported genes TaMBF1c and TaHSFA6e . Gene Ontology enrichment indicated these genes are involved in "response to heat", "protein folding", and "response to reactive oxygen species". Integrating DEGs, core genes, and prior data, we selected three candidate genes: TaCPD-1 , TaHSF-1 , and TaBI-1 , with TaMBF1c employed as the positive control in the experiment. Yeast assays showed that TaMBF1c , TaBI-1 , and TaCPD-1 overexpression enhanced thermotolerance under 50 ℃. Transient expression in wheat via nano-carriers confirmed that TaMBF1 c and TaBI-1 overexpression improve heat tolerance under 42 ℃, outperforming wild-type and control plants. Our results demonstrate that TaMBF1c and TaBI-1 enhance thermotolerance in both heterologous and native systems. This study provides a theoretical basis for elucidating the molecular mechanisms of heat stress responses in wheat and for cultivating heat-tolerant wheat varieties.